The Impact of Making Proud Choices! on Youth's Sexual Health Attitudes, Knowledge, and Behaviors.

PURPOSE: To estimate the effect of the Making Proud Choices! (MPC) curriculum, one of the most popular evidence-based teen pregnancy prevention curricula being implemented nationwide. METHODS: A cluster randomized trial was used to assign high schools to receive the MPC program or to serve as a standard care comparison group. The study took place in 15 schools in four cities in the United States. A total of 31 clusters and over 2,800 youth (predominantly in grades 9 and 10) were assigned to condition between 2016 and 2019. Student surveys administered at baseline and approximately 9 months after baseline were used to estimate the effects of MPC on risk and protective factors (knowledge, attitudes, self-efficacy) as well as sexual behavior outcomes. RESULTS: The evaluation showed several large, statistically significant, and favorable impacts on nearly all of the risk and protective factors for risky sexual behavior. In addition, the study found a favorable effect of MPC on reducing the number of episodes of sex in the past 3 months. DISCUSSION: The evaluation showed that MPC, as it is commonly implemented today, has evidence of effectiveness on improving both risk and protective factors, and sexual behavior outcomes. Based on these findings, MPC will continue to be seen as an evidence-based teen pregnancy prevention program for implementation through federal grant programs.

Effects of the Nurturing Parenting Program Nurturing Skills for Families on child safety and permanency.

BACKGROUND: Many child welfare agencies use the Nurturing Parenting Program Nurturing Skills for Families (NPP) program to strengthen parenting skills. NPP includes a flexible sequence of lessons that are tailored to meet each family's needs. OBJECTIVE: This study estimated the effects of NPP on child safety and permanency outcomes using a quasi-experimental design. PARTICIPANTS AND SETTING: This study included 1102 children in Arizona whose families were referred to NPP between 2018 and 2020 (treatment group) and 6845 children in Arizona whose families were referred to other in-home family preservation services during the same period (comparison group). METHODS: Outcomes were based on child welfare administrative data. The study estimated (1) the effects of being referred to NPP (regardless of a family's level of participation) and (2) the effects of completing NPP. Baseline equivalence was established for each analysis. Impacts were calculated by comparing regression-adjusted differences between the study groups. RESULTS: The study found no evidence of impacts of being referred to NPP. However, children whose families completed NPP were less likely to experience an investigation (ES = -0.28; p = 0.03) or substantiated investigation (ES = -0.66; p = 0.03) 4 months after the service referral, and less likely to experience a removal 16 months later (ES = -0.70; p = 0.00). CONCLUSIONS: NPP had favorable effects on child welfare outcomes when families completed the program. Additional research is necessary to understand the supports that enable families to complete NPP and the specific components that are particularly effective.

Characterizing cell interactions at scale with made-to-order droplet ensembles (MODEs).

Cell-cell interactions are important to numerous biological systems, including tissue microenvironments, the immune system, and cancer. However, current methods for studying cell combinations and interactions are limited in scalability, allowing just hundreds to thousands of multicell assays per experiment; this limited throughput makes it difficult to characterize interactions at biologically relevant scales. Here, we describe a paradigm in cell interaction profiling that allows accurate grouping of cells and characterization of their interactions for tens to hundreds of thousands of combinations. Our approach leverages high-throughput droplet microfluidics to construct multicellular combinations in a deterministic process that allows inclusion of programmed reagent mixtures and beads. The combination droplets are compatible with common manipulation and measurement techniques, including imaging, barcode-based genomics, and sorting. We demonstrate the approach by using it to enrich for chimeric antigen receptor (CAR)-T cells that activate upon incubation with target cells, a bottleneck in the therapeutic T cell engineering pipeline. The speed and control of our approach should enable valuable cell interaction studies.

Teen Pregnancy Prevention Evaluation Technical Assistance: A Case Study.

PURPOSE: This case study discusses Mathematica's experience providing large-scale evaluation technical assistance (ETA) to 65 grantees across two cohorts of Teen Pregnancy Prevention (TPP) Program grants. The grantees were required to conduct rigorous evaluations with specific evaluation benchmarks. This case study provides an overview of the TPP grant program, the evaluation requirements, the ETA provider, and other key stakeholders and the ETA provided to the grantees. Finally, it discusses the successes, challenges, and lessons learned from the effort. CONCLUSION: One important lesson learned is that there are two related evaluation features, strong counterfactuals and insufficient target sample sizes, that funders should attend to prior to selecting awardees because they are not easy to change through ETA. In addition, if focused on particular outcomes (for TPP, the goal was to improve sexual behavior outcomes), the funder should prioritize studies with an opportunity to observe differences in these outcomes across conditions; several TPP grantees served young populations, and sexual behavior outcomes were not observed or were rare, limiting the opportunity to observe impacts. Unless funders are attentive to weaning out evaluations with critical limitations during the funding process, requiring grantees to conduct impact evaluations supported by ETA might unintentionally foster internally valid, yet underpowered studies that show nonsignificant program impacts. The TPP funder was able to overcome some of the limitations of the grantee evaluations by funding additional evidence-building activities, including federally led evaluations and a large meta-analysis of the effort, as part of a broader learning agenda.

The impact of the Regional Partnership Grant program on adult recovery and well-being, and child safety, permanency, and well-being outcomes.

BACKGROUND: Adult substance use contributes to high rates of child maltreatment, adverse parenting, and poor child well-being outcomes. The Children's Bureau in the Administration for Children and Families, U.S. Department of Health and Human Services funded the Regional Partnership Grant (RPG) program to improve safety, well-being, and permanency outcomes for children at risk of or in out-of-home placements because of caregivers' substance use. OBJECTIVE: This study estimates the effect of the RPG program on adult parenting and recovery outcomes, as well as child safety, permanency, and well-being outcomes. PARTICIPANTS AND SETTING: This study included samples from three RPG projects funded in 2014. A total of 437 families were included in the evaluation: 198 enrolled in RPG and 239 in the comparison group. METHODS: Outcomes included both administrative data and standardized instruments collected at program entry and exit. Impacts were calculated by comparing regression-adjusted differences between RPG participants and a well-matched comparison group. RESULTS: The RPG program showed two favorable and statistically significant impacts on adult outcomes: reduced trauma symptoms (ES = -0.25, p = 0.02), and depressive symptoms (ES = -0.25, p = 0.03). In addition, children enrolled in RPG had reduced behavior problems (ES = -0.31, p = 0.02) and reduced executive functioning deficits (ES = -0.31, p = 0.04). There were no significant impacts observed for child safety and permanency outcomes. CONCLUSIONS: The RPG approach has produced several favorable impacts on key mental health needs of caregivers and children, establishing foundational evidence of program effectiveness. Additional evaluation research is necessary to understand the effectiveness on additional outcomes with a larger, more generalizable sample.

Simple, Affordable, and Modular Patterning of Cells using DNA.

The relative positioning of cells is a key feature of the microenvironment that organizes cell-cell interactions. To study the interactions between cells of the same or different type, micropatterning techniques have proved useful. DNA Programmed Assembly of Cells (DPAC) is a micropatterning technique that targets the adhesion of cells to a substrate or other cells using DNA hybridization. The most basic operations in DPAC begin with decorating cell membranes with lipid-modified oligonucleotides, then flowing them over a substrate that has been patterned with complementary DNA sequences. Cells adhere selectively to the substrate only where they find a complementary DNA sequence. Non-adherent cells are washed away, revealing a pattern of adherent cells. Additional operations include further rounds of cell-substrate or cell-cell adhesion, as well as transferring the patterns formed by DPAC to an embedding hydrogel for long-term culture. Previously, methods for patterning oligonucleotides on surfaces and decorating cells with DNA sequences required specialized equipment and custom DNA synthesis, respectively. We report an updated version of the protocol, utilizing an inexpensive benchtop photolithography setup and commercially available cholesterol modified oligonucleotides (CMOs) deployed using a modular format. CMO-labeled cells adhere with high efficiency to DNA-patterned substrates. This approach can be used to pattern multiple cell types at once with high precision and to create arrays of microtissues embedded within an extracellular matrix. Advantages of this method include its high resolution, ability to embed cells into a three-dimensional microenvironment without disrupting the micropattern, and flexibility in patterning any cell type.

High diversity droplet microfluidic libraries generated with a commercial liquid spotter.

Droplet libraries consisting of many reagents encapsulated in separate droplets are necessary for applications of microfluidics, including combinatorial chemical synthesis, DNA-encoded libraries, and massively multiplexed PCR. However, existing approaches for generating them are laborious and impractical. Here, we describe an automated approach using a commercial array spotter. The approach can controllably emulsify hundreds of different reagents in a fraction of the time of manual operation of a microfluidic device, and without any user intervention. We demonstrate that the droplets produced by the spotter are similarly uniform to those produced by microfluidics and automate the generation of a ~ 2 mL emulsion containing 192 different reagents in ~ 4 h. The ease with which it can generate high diversity droplet libraries should make combinatorial applications more feasible in droplet microfluidics. Moreover, the instrument serves as an automated droplet generator, allowing execution of droplet reactions without microfluidic expertise.

An Oil-Free Picodrop Bioassay Platform for Synthetic Biology.

Droplet microfluidics enables massively-parallel analysis of single cells, biomolecules, and chemicals, making it valuable for high-throughput screens. However, many hydrophobic analytes are soluble in carrier oils, preventing their quantitative analysis with the method. We apply Printed Droplet Microfluidics to construct defined reactions with chemicals and cells incubated under air on an open array. The method interfaces with most bioanalytical tools and retains hydrophobic compounds in compartmentalized reactors, allowing their quantitation.

Development and evaluation of a new diffusive gradients in thin-films technique for measuring organotin compounds in coastal sediment pore water.

Organotins present a toxicological risk to biota in the aquatic environment. Understanding the behaviour of these compounds in sediment is challenging, with sophisticated analytical techniques required for their measurement. We investigated the use of silica-bound sorbents for diffusive gradients in thin-films (DGT) adsorption gels to pre-concentrate five organotins (monobutlytin (MBT), dibutyltin (DBT), tributyltin (TBT), diphenyltin (DPhT), triphenyltin (TPhT)) found frequently in coastal sediment. C8 sorbent showed optimum performance in uptake and recovery of organotins for pH and ionic strength ranges typical of coastal waters. Recoveries from adsorption gels deployed in filtered sea water were MBT = 123 ± 20%, DBT = 75 ± 12%, TBT = 81 ± 16%, DPhT = 72 ± 30%, TPhT = 58 ± 10% respectively. Devices were used to investigate DGT fluxes and pore water concentrations of organotins in coastal sediment collected from a contaminated site. DGT fluxes measured in sediment cores for the five organotins ranged between 4.3 × 10-8 and 1.6 ×10-5ngcm2s-1. The depletion of organotin species within pore waters at the interface with DGT devices was measured over a series of deployment times (2, 7, 14, 21 and 28 days) and provided estimates of the concentration of organotins in pore waters at Langstone Harbour, UK, prior to depletion by the DGT device and information on their spatial heterogeneity. The novel in situ DGT device developed can pre-concentrate organotins from pore waters in coastal sediment core samples and allows their detection at low environmental concentrations using conventional gas chromatographic/mass spectrometric instrumentation. Use of the DGT device overcomes many problems associated with the conventional pore water sampling of organotins. Our preliminary data suggests it has potential in the future to be a useful tool in investigating the environmental fate of these pollutants. The use of the C8 gel will also allow for the simultaneous sequestration of other semi- and non-polar analytes present in the pore water.

Printed droplet microfluidics for on demand dispensing of picoliter droplets and cells.

Although the elementary unit of biology is the cell, high-throughput methods for the microscale manipulation of cells and reagents are limited. The existing options either are slow, lack single-cell specificity, or use fluid volumes out of scale with those of cells. Here we present printed droplet microfluidics, a technology to dispense picoliter droplets and cells with deterministic control. The core technology is a fluorescence-activated droplet sorter coupled to a specialized substrate that together act as a picoliter droplet and single-cell printer, enabling high-throughput generation of intricate arrays of droplets, cells, and microparticles. Printed droplet microfluidics provides a programmable and robust technology to construct arrays of defined cell and reagent combinations and to integrate multiple measurement modalities together in a single assay.

A simple, low cost GC/MS method for the sub-nanogram per litre measurement of organotins in coastal water.

Tributyltin (TBT) is a legacy pollutant in the aquatic environment, predominantly from its use in anti-foulant paints and is listed as a priority hazardous substance in the European Union's Water Framework Directive (WFD). Measuring low concentrations of TBT and other organotins (e.g. monobutyltin (MBT), dibutyltin (DBT), diphenyltin (DPhT) and triphenyltin (TPhT)) at sub ng/L concentrations in coastal waters using standard laboratory instrumentation is very challenging. Conventional, low injection volume gas chromatography/mass spectrometry (GC/MS) combined with liquid-liquid extraction typically achieves limits of detection for TBT ∼10 ng L(-1). We describe a simple, programmed temperature vaporisation-large injection volume (50 µL), GC/MS selected ion monitoring method for measuring DBT, TBT, DPhT and TPhT in coastal waters at lower concentrations. Quantification of MBT was not possible using these injection volumes but was achieved using a 10 µL injection volume together with a reduced injection speed. This new approach offers: •When using a 50 µL injection, limits of detection = 0.70 ng L(-1) and limits of quantification = 2.1 ng L(-1) for TBT were achieved in derivatised standards.•Recoveries of TBT and TPhT from coastal water >97%.•Time consuming, off-line sample pre-concentration methods are unnecessary.

Multicolor Fluorescence Detection for Droplet Microfluidics Using Optical Fibers.

Fluorescence assays are the most common readouts used in droplet microfluidics due to their bright signals and fast time response. Applications such as multiplex assays, enzyme evolution, and molecular biology enhanced cell sorting require the detection of two or more colors of fluorescence. Standard multicolor detection systems that couple free space lasers to epifluorescence microscopes are bulky, expensive, and difficult to maintain. In this paper, we describe a scheme to perform multicolor detection by exciting discrete regions of a microfluidic channel with lasers coupled to optical fibers. Emitted light is collected by an optical fiber coupled to a single photodetector. Because the excitation occurs at different spatial locations, the identity of emitted light can be encoded as a temporal shift, eliminating the need for more complicated light filtering schemes. The system has been used to detect droplet populations containing four unique combinations of dyes and to detect sub-nanomolar concentrations of fluorescein.

Compact and modular multicolour fluorescence detector for droplet microfluidics.

Multicolour fluorescence detection is often necessary in droplet microfluidics, but typical detection systems are complex, bulky, and expensive. We present a compact and modular detection system capable of sub-nanomolar sensitivity utilizing an optical fibre array to encode spectral information recorded by a single photodetector.

Double Emulsion Generation Using a Polydimethylsiloxane (PDMS) Co-axial Flow Focus Device.

Double emulsions are useful in a number of biological and industrial applications in which it is important to have an aqueous carrier fluid. This paper presents a polydimethylsiloxane (PDMS) microfluidic device capable of generating water/oil/water double emulsions using a coaxial flow focusing geometry that can be fabricated entirely using soft lithography. Similar to emulsion devices using glass capillaries, double emulsions can be formed in channels with uniform wettability and with dimensions much smaller than the channel sizes. Three dimensional flow focusing geometry is achieved by casting a pair of PDMS slabs using two layer soft lithography, then mating the slabs together in a clamshell configuration. Complementary locking features molded into the PDMS slabs enable the accurate registration of features on each of the slab surfaces. Device testing demonstrates formation of double emulsions from 14 µm to 50 µm in diameter while using large channels that are robust against fouling and clogging.

Assessing environmentally significant effects: a better strength-of-evidence than a single P value?

Interpreting a P value from a traditional nil hypothesis test as a strength-of-evidence for the existence of an environmentally important difference between two populations of continuous variables (e.g. a chemical concentration) has become commonplace. Yet, there is substantial literature, in many disciplines, that faults this practice. In particular, the hypothesis tested is virtually guaranteed to be false, with the result that P depends far too heavily on the number of samples collected (the 'sample size'). The end result is a swinging burden-of-proof (permissive at low sample size but precautionary at large sample size). We propose that these tests be reinterpreted as direction detectors (as has been proposed by others, starting from 1960) and that the test's procedure be performed simultaneously with two types of equivalence tests (one testing that the difference that does exist is contained within an interval of indifference, the other testing that it is beyond that interval-also known as bioequivalence testing). This gives rise to a strength-of-evidence procedure that lends itself to a simple confidence interval interpretation. It is accompanied by a strength-of-evidence matrix that has many desirable features: not only a strong/moderate/dubious/weak categorisation of the results, but also recommendations about the desirability of collecting further data to strengthen findings.

MP4, a vasodilatory PEGylated hemoglobin.

A vasodilatory hemoglobin (Hb)-based O(2) carrier (HBOC) has been developed by surface conjugation polyethylene glycol to tetrameric human Hb (MP4, Sangart, San Diego). Because the NO-binding kinetics of MP4 are similar to vasoconstrictive HBOCs, we propose that the decoupling of NO scavenging from vascular response is a consequence of MP4's high O(2) affinity (p50 = 5 mmHg) and unique surface chemistry. The release of ATP from erythrocytes is vasodilatory and the application of a high O(2) affinity HBOC minimizes ATP interference with intravascular ATP signaling. A second potential mechanism of action for MP4 involves the surface conjugation of polyethylene glycol (PEG) to tetrameric human Hb. It has been shown that the addition of unconjugated high molecular weight (Mw) PEG to cultured lung endothelial cells causes an immediate and significant reduction in endothelial permeability; an effect opposite to that of endothelial agonists such as cell-free Hb. It appears that some of the benefits of the PEG-endothelium interaction are carried onto molecules such as PEGylated Hb and PEGylated albumin, as demonstrated by favorable hemodynamic responses in vivo. PEGylation of ß93 cysteine residues, as in MP4, has also been reported to increase the nitrite reductase activity of Hb and enhance conversion of endogenous nitrite to bioactive NO.